The light chain component of the Ig protein is encoded by 2 separate loci, Igκ and Igλ. The proportion of antibodies containing κ or λ light chains varies considerably between different species (1-3), e.g., in mice the κ:λ ratio is 95:5, compared to 60:40 in humans. Two models have evolved to account for this apparent bias in the expression of κ in the mouse. First, from the observation that murine Igλ-producing myelomas have rearranged κ light chain genes, and that Igκ producing cells have the λ light chain locus in germline configuration, it was proposed that κ rearrangement must occur before λ rearrangement can commence (4, 5). In the human situation, however, while almost all λ producing cells have both κ alleles rearranged, the proportion of κ and λ producing cells are similar (4). The second proposal is that κ and λ loci are both available for rearrangement at the same time, but the mouse κ locus is more efficient at engaging the rearrangement process (6). The occasional finding of cells with rearranged λ and the κ locus in germline configuration may support this (5, 7, 8). The influence of antigen selection on the biased κ:λ ratio is discounted by the finding that the ratio is similar in fetal liver and in cells that have not encountered antigen (9-13).
Light chain V-J rearrangement occurs at the transition from pre B-II to immature B cells, where the surrogate light chain associated with membrane Igμ is replaced by κ or λ light chain (14). Although the timing of light chain rearrangement is essentially defined, the processes which activate light chain locus rearrangement are not fully understood. From locus silencing experiments, it became clear that κ rearrangement is not a prerequisite for λ recombination (15). Indeed, κ and λ rearrangements are independent events (16), the activation of which may be affected by differences in the strength of the respective enhancers. A region believed to be important in the regulation of the accessibility of the human λ locus has been identified about 10 Kb downstream of Cλ7 (17, 18). Functional comparisons in reporter gene assays identified a core enhancer region that is flanked by elements which can drastically reduce enhancer activity in pre-B cells (17). Although transfection studies showed that the κ and λ 3′ enhancer regions appear to be functionally equivalent, other (functional) sequences flanking the core enhancer motifs are remarkably dissimilar. Targeted deletion of the κ 3′ enhancer in transgenic mice showed that this region is not essential for κ locus rearrangement and expression but is required to establish the κ:λ ratio (19).
The human Igλ locus on chromosome 22q11.2 is 1.1 Mb in size and typically contains 70 Vλ genes and 7 Jλ-Cλ gene segments (20, 21 and references therein). About half of the Vλ genes are regarded as functional and Jλ-Cλ1, 2, 3 and 7 are active. The Vλ genes are organized in 3 clusters which contain distinct V gene family groups. There are 10 Vλ gene families, with the largest VλIII being represented by 23 members. In human peripheral blood lymphocytes, the most J-C proximal V gene segments in cluster A, from families I, II and III, are preferentially rearranged, with the contribution of the 2a2 Vλ segment (2-14 in the new nomenclature (22) being unusually high (23). All λ gene segments have the same polarity which allows deletional rearrangement (24). Sequence diversity of the Igλ repertoire is provided mainly by Vλ-Jλ combination. Additional CDR3 diversity due to N (nonencoded)- or P (palindromic)-nucleotide additions at the V to J junction, although not as extensive as seen in IgH rearrangement, seems to be much more frequently used in humans than in mice (25, 26, 27, 28), where the TdT (terminal deoxyribonucleotide transferase) activity is down-regulated at the time of light chain rearrangement.
It has been shown that human Ig can be produced in transgenic mice carrying human Ig genes on miniloci or yeast artificial chromosomes (YACs) (58, 59, 60, 61, 62) and that silencing of the endogenous mouse heavy and κ loci enhances human antibody production in such transgenic animals. However, in all such mice reported to date, only the human κ light chain genes have been incorporated and there have been no reports of the human λ light chain locus being integrated into transgenic mice. Therefore, until the present invention, no λ-containing human antibodies have been made from transgenic mice, nor has there been any information on the expressibility of human λ genes in such animals or on the relative contributions of human κ and λ in mice carrying both transgenic human loci. Thus it was not known whether λ-transgenic mice would be suitable for the production of human antibodies.